| BackgroundBurn patients are at high risk for local and systemic infections.Although infection control programme has been performed in most burn centres and hospitals,the incidence of NI remains high.Infection is one of the main causes of death in burn patients,and sepsis is the most serious manifestation of infection,which is the most important cause of failure of burn treatment,and has a decisive influence on the clinical outcome of burn treatment.The incidence rate of sepsis is increasing year by year worldwide,and sepsis is considered to be one of the leading causes of death of critically ill patients.The concept of SEPSIS 1.0 was proposed in the early 1990s.Despite the subsequent proposal of SEPSIS2.0 years later,sepsis was still defined as the systemic inflammatory response(SIRS)to infection.SEPSIS1.0 and SEPSIS2.0 in terms of SIRS have been used as a clinical standard for judging whether or not patients suffer from sepsis.Furthermore,SIRS is considered to be the chief culprit in the induction of multiple organ dysfunction syndrome(MODS)in sepsis.In 2016,however,the concept of SIRS has been eliminated from SEPSIS3.0.This updated definition implies that SIRS is no longer necessary for the diagnosis of sepsis,opposing the previous clinical diagnostic criteria.However,years of basic and clinical studies have shown that sepsis is caused by a complex interaction between the host immune system and pathogenic microorganisms,in which cytokines play an important role in the imbalance of immune inflammatory response.For instance,"cytokine storm" is considered to be the major driver of morbidity and mortality for severe COVID-19 patients who suffer from septic shock.Therefore,it is very important to explore the potential role of inflammatory factors in the pathophysiological process of sepsis.Similar to TNF,Interleukin-1β(IL-1β),produced primarily by monocytes and macrophages in response to endotoxin,is another inflammatory factor of SIRS and plays an irreplaceable role in sepsis.Previous studies mostly focused on the negative role of excessive IL-1 release in sepsis,and improved the prognosis of sepsis by antagonizing inflammatory mediators,but the results were not the same.Some studies have previously revealed that prophylactic administration of IL-1 can protect mice from lethal Gram-negative infection.A recent research has shown that IL-1 receptor antagonist treatment aggravates staphylococcal septic arthritis and sepsis in mice[10].Additionally,IL-1β contributes to macrophage recruitment and Streptococcus pneumoniae clearance.All these findings suggest that IL-1β may improve the prognosis of sepsis by initiating innate immunity in sepsis.However,the immune-regulatory mechanism of IL-1 remains largely unknown.Regulatory dendritic cells(DCregs)play a key role in sepsis.DCregs can attenuate the excessive production of inflammatory cytokines in the early stage of patient trauma,prevent the excessive inflammatory response from causing tissue damage,and regulate the balance between pro-inflammatory and anti-inflammatory response.DCregs play an important role in the regulation of immune balance.When negative immune regulation is dominant,the body’s ability to inhibit excessive inflammatory response will be strengthened,so as to effectively prevent the immune suppression state at the late stage of sepsis,multiple organ dysfunction syndrome and even death caused by excessive inflammatory response.The change of DCregs may be a key factor for the body to resist potential attack.The increase of the proportion of DCregs in the later stage of sepsis can inhibit the degree of excessive inflammatory response when potential attack occurs,so that the body can be in a state of immune balance.Previous studies have reported that IL-1β accelerates hematopoietic recovery in the bone marrow(BM).Recent work in our laboratory has demonstrated that High Mobility Group Protein 1(HMGB1)improves the prognosis of sepsis by boosting differentiation of bone marrow cells(BMCs)into regulatory dendritic cells(DCs).Thus,we hypothesize that IL-1β can enhance innate immunity by stimulating local proliferation and differentiation of BMCs into CD11c-CD45RBhigh DCs in both immune and non-immune organs,thereby leading to sufficient bacterial eradication and the alleviation of sepsis.In conclusion,sepsis,the most severe manifestation of infection,is the main cause of death in severe burn patients.Inflammatory cytokines and dendritic cells play an important role in the occurrence and development of sepsis,and their research is of positive significance for the knowledge,understanding and development of possible effective treatment measures for sepsis.Objectives1.To investigate the influence of infection on the clinical outcome of burn patients,and to understand the progression of sepsis after burn infection.2.To explore the protective effect of IL-1β on sepsis mice.3.To study the effect of IL-1β on endogenous IL-1β release in sepsis mice.4.To study the effect of IL-1β on the proliferation and differentiation of bone marrow and liver immune cells in sepsis mice.5.To study the effect of adoptive transfusion of bone marrow and liver immune cells and regulation of dendritic cells on the prognosis of sepsis mice.Methods1.Use competing risk and multistate model to estimate the impact of nosocomial infection on length of stay and mortality in burn patients.The proportion of patients who developed sepsis after burn infection was observed.2.CLP sepsis model was established.Mice were given different doses of IL-1β.The survival rate of mice 14 days after CLP operation was observed to explore the effects of different concentrations of IL-1β on the prognosis of mice with CLP sepsis.Sepsis mice were given the same dose(50μg/kg)of IL-1β at different time points after CLP operation.The survival rate of sepsis mice at 14 days was observed,and the effect of IL-1β at different time points on the prognosis of CLP sepsis model mice was explored.3.To establish a mouse model of escherichia coli induced sepsis.To observe the survival rate of sepsis mice which were given different doses of IL-1β 14 days after operation.To explore the effect of different concentrations of IL-1β on the prognosis of escherichia coli induced sepsis model mice.Sepsis mice were given the same dose of IL-1β(25μg/kg)at different time points,and the 14-day survival rate of mice was observed.To explore the effect of IL-1β administration at different time points on the prognosis of escherichia coli induced sepsis model mice.4.The serum IL-1β level was observed at 0.1,12,24,48,72 h after intraperitoneal injection of the CLP sepsis model.The serum IL-1β level was observed at 0.1,1,3,24,24 h after intraperitoneal injection of the escherichia coli sepsis model.5.After the establishment of CLP sepsis model,different concentrations of IL-1βwere immediately injected intraperitoneally to observe the survival rate of sepsis mice at different time points.The number of immune cells in liver and bone marrow and the proportion of regulatory dendritic cells(CD1lc-CD45RBhigh)in sepsis mice were isolated and detected at 0,3,6,9,14 days after CLP operation.The expression of CD40,CD80,CD86 and I-A/E and the secretion of IL-10 and IL-12 in CD11c-CD45RBhighcells were detected by flow cytometry and ELISA.To investigate whether IL-1β can affect the pathophysiological process of sepsis by inducing the proliferation of bone marrow and liver immune cells and differentiating into CD11c-CD45RBhighDCs in sepsis mice.6.BMC,liver immune cells and CD11c-CD45RBhigh DC were isolated from CLP sepsis model mice 9 days after intrabitoneal injection of IL-1β(50μg/kg).Different numbers of immune cells were adopted and returned to CLP sepsis model mice by intraperitoneal injection.To observe the survival rate of sepsis mice at different time points,and whether there was a dose-dependent relationship between the survival rate of sepsis mice and the number of adoptive immune cells.To investigate whether IL-1β induces CD1lc-CD45RBhigh DC differentiation and improves immune function by stimulating the proliferation of bone marrow and liver immune cells in CLP sepsis mice,thus affecting the prognosis of CLP sepsis mice.7.Statistical analysis:Data were expressed as mean ± standard deviation(SD).Experimental data were analyzed with Social Sciences(SPSS),version18.0(SPSS,Chicago,Inc.,IL)and results were analyzed by t test,one-way variance(ANOVA)test with Bonferroni ’s post hoc test for pairwise comparisons and log rank test.p values below 0.05 was considered significant.Results1.INosocomial infection was closely associated with increased mortality and prolonged hospital stay in burn patients,and SOFA>2 in 48%of burn patients progressed to sepsis.2.IL-1β improves the survival rate of CLP sepsis model mice:after intraperitoneal injection of 50μg/kg IL-1β immediately after CLP,the survival rate at 14 days was 80%,which was significantly higher than that of the control group treated with normal saline(the survival rate at 14 days was 10%).Mice injected with IL-1β at doses of 5-12.5 μg/kg or 100-125 μg/kg had similar survival to the control group,but there was no significant survival advantage.Sepsis mortality did not decrease when IL-1β was injected at a dose less than 5μg/kg,but significantly increased when IL-1β was injected at a dose greater than 500μg/kg.After intraperitoneal injection of IL-1β(50μg/kg)CLP 1h before surgery and immediately after surgery,the 14-day survival rate was 90%and 70%,respectively,significantly higher than that of the control group(14-day survival rate was 10%).There was no significant improvement in survival after injection of IL-1β at 1 and 6h after CLP.IL-1β was injected at intervals of 12 to 48 hours,and there was no observed difference in survival compared with the control group.The results showed that IL-1β could significantly improve the survival rate of CLP sepsis model mice.The optimal dose of IL-1β was 50μg/kg,and the optimal time of administration was 1h before and immediately after CLP.IL-1β improves survival in mice with E.coli induced sepsis:the 14-day survival rate of mice with sepsis after intraperitoneal injection of 25μg/kg IL-1βwas 80%,compared with 0%in the control group.The survival rate of sepsis mice treated with 12.5 μg/kg IL-1β or 50μg/kg IL-1β was significantly higher than that of the control group.Sepsis mortality did not decrease when the dose of IL-1βwas less than 2.5μg/kg,and sepsis mice did not survive when the dose of IL-1βwas higher than 250 μg/kg.IL-1β(25μg/kg)was administered 1h and immediately before escherichia coli induced sepsis,and the 14-day survival rate was 80%and 50%,respectively.Injection of IL-1β 1h after E.coli infection did not improve survival,and no difference in survival was observed between the control group and the 6-12h interval.The results showed that IL-1β could significantly improve the survival rate of escherichia coli induced sepsis model mice,and had protective effect on escherichia coli fatal infection.The optimal dose of IL-1β was 25μg/kg,and the optimal time of administration was 1h before operation and immediately after operation.3.IL-1β inhibited the release of IL-1β in the early stage of sepsis:After intraperitoneal injection of IL-1β immediately after CLP and fatal E.coli infection,the level of serum IL-1β in mice gradually decreased,while the level of serum IL-1β in CLP and EScherichia coli infection control mice gradually increased in saline treatment.The level of IL-1β in the CLP group was similar to that in the control group at 48h,and the level of IL-1β in the fatal e.coli group at 3h was similar to that in the control group.The results showed that IL-1βinhibited the release of IL-1β in the early stage of sepsis.4.IL-1β stimulation increased the number of bone marrow and liver immune cells in CLP sepsis mice:The survival rate of CLP sepsis mice treated with IL-1β was higher than that of untreated sepsis mice,and a dose-dependent reduction in mortality was observed with injection of 50 μg/kg IL-1β compared with intraperitoneal injection of 12.5 or 25 μg/kg IL-1β.After intraperitoneal injection of 12.5-50 μg/kg IL-1β immediately after CLP,the number of BMC and liver immune cells increased gradually from 3 days to 9 days after CLP,and on day 14 after CLP,the number of BMC and liver immune cells was close to the normal level,and there was no significant difference among the groups.5.IL-1β stimulation increased the number and percentage of CD11c-CD45RBhighDC in bone marrow and liver of CLP sepsis mice:The number and proportion of CD11c-CD45RBhighDC in bone marrow and liver immune cells were significantly increased 3 to 9 days after intraperitoneal injection of 50 μg/kg IL-1β in CLP sepsis mice.Subsequently,9 to 14 days after CLP,the number and percentage of CD11c-CD45RBhighDC gradually approached that of the sham treatment group.The phenotype and function of CD11c-CD45RBhighDC in BM and liver were similar to that in spleen,with low expression of CD40,CD80,CD86 and I-A/E and high secretion of IL-10.The results suggest that IL-1β induces the differentiation of bone marrow and liver immune cells into CD11c-CD45RBhighDC,which may play a key role in the pathophysiological process of sepsis.6.Beneficial immunomodulation of septic BMCs and liver immune cells treated by IL-1β in vivo:A single intraperitoneal injection of BMCs or CD11c-CD45RBhighDCs in BM 9 days post-CLP,isolated from CLP mice with IL-1β(50μg/kg)stimulation,could reduce the mortality of septic mice in a dose-dependent manner.Surprisingly,a single intraperitoneal injection of liver immune cells or CD11c-CD45RBhighDCs in liver 9 days post-CLP,isolated from CLP mice with IL-1β(50 μg/kg)stimulation,could also reduce the mortality of septic mice in a dose-dependent manner.Based on these data,the numbers of BMCs and liver immune cells were increased,and the immune function of these cells in CLP mice with IL-1β treatment was enhanced remarkably.ConclusionInfection was closely associated with prolonged hospitalization and increased mortality in burn patients,and nearly half of those with burn infection progressed to sepsis.IL-1β has a protective effect on sepsis by stimulating the proliferation of bone marrow and liver immune cells in sepsis mice,inducing them to differentiate into CD11c-CD45RBhighDC,and coordinating the redistribution of immune cells in immune and non-immune organs,thus improving the immune function of sepsis. |
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